Babesiosis is a tick-borne, intraerythrocytic protozoan infection, caused by members of the Babesia genus. Transfusion-transmitted babesiosis (TTB) is well described: 162 cases of TTB have been reported in the US alone, the overwhelming majority caused by Babesia microti, which is widely endemic in the Northeastern and upper Midwestern US. An increase in naturally-acquired and TTB, has resulted in B. microti's designation as the foremost infectious risk to blood safety in the US for which an effective screening strategy is currently unavailable. Uncertainty surrounding the epidemiology, transmissibility and immunopathogenesis of B. microti remains an obstacle to development of a viable mitigation strategy. Recent evidence suggests that transfusion transmission may be caused disproportionately by a subset of chronically parasitemic blood donors. This has prompted our aims: 1) to determine the proportions of B. microti seropositive donors that develop chronic versus transient/resolved infections, and 2) to build a specimen repository composed of longitudinal samples obtained from B. microti sero- and or PCR-positive blood donors for future characterization in order to further our understanding of the mechanisms underlying chronic infection. We plan to leverage two independently funded, planned studies which will screen 20,000 blood donations in New York from July to September in 2012 (SBIR) and 2013 (IND), using a prototype highly sensitive and specific ELISA to detect antibodies against B. microti. Seroreactive samples will be subjected to supplementary testing using a novel, highly sensitive real- time PCR-based assay in addition to IFA and peripheral blood smear examination (PBS). Under the R21, we plan to enroll these seroreactive donors (n=100) and to conduct serial blood sampling and clinical symptom/risk factor questionnaire administration over 1 year of follow-up. At each follow-up visit, we will perform ELISA, PCR, IFA and PBS testing, in addition to repeated administration of the clinical questionnaire. We will also enroll donors that have previously been implicated in cases of TTB (n=10) and donors that have been deferred following their self-reporting a history of babesiosis. After testing, residual whole blood and plasma from each follow-up visit will be frozen and archived in a repository for future characterization. Correlation of the index and follow-up results (ELISA and PCR) will enable infectious-risk categorization into: chronic infection (ELISA+/PCR+), transient infection (index PCR+/follow-up PCR -) and remote exposure (index and follow-up PCR-). This categorization informs choice of screening methodology (serology vs. PCR) and policy of deferral and reinstatement. We intend to use the sample repository for a future submission to determine the genetic and immunologic determinants of chronic vs. transient infection.